The cloned, functional mouse pituitary adenocarcinoma cell line, AtT-20 is grown in tissue culture, the D-16 clone in large-volume roller culture. Using bioassays for ACTH and MSH, radioimmunoassays for ACTH, alpha-MSH and beta-MSH, we have shown this cell secretes two ACTH moieties and an MSH as well. Synthesis of ACTH is specifically inhibited by physiological concentrations of glucocorticoids, but not by other steroid hormones. The steroids bind to a cytoplasmic receptor protein whose binding affinity for various steroids parallels their inhibitory effects on ACTH synthesis. The steroid-receptor complex translocates into the nucleus, where it binds to nucleoprotein. The AtT-20 cell line has a temperature-dependent steroid membrane transport mechanism, with a component to which steroids bind prior to binding to the cytoplasmic receptor molecule. The AtT-20 cell also produces a C-type virus particle which appears to be a MuLV by indirect immunofluorescence and XC syncytial cell assay. The human EPO malignant melanoma clonal cell line isolated in this laboratory, secretes "ectopic" ACTH. Synthesis of ACTH is not inhibited by glucocorticoids, and the cells lack a cytoplasmic receptor for glucocorticoids, although the isolated EPO nucleus will take up and bind steroid complexed with the AtT-20 cytosol receptor.